维甲酸诱导P19胚胎癌细胞向神经元分化过程中分化相关因子的表达  

Expression of Differentiation Related Factors in the Process of Retinoid Acid Inducing P19 Embryonic Carcinoma Cell Differentiation into Neuron Cells

作　　者：张文迪[1] 付溪[1] 高金枝[1] 谢雪梅[1] 罗小平[1] 

机构地区：[1]华中科技大学同济医学院附属同济医院儿科,武汉430030

出　　处：《神经损伤与功能重建》2013年第3期171-176,共6页Neural Injury and Functional Reconstruction

基　　金：国家重点基础研究发展项目(973计划)(No.2005CB5225007);卫生部临床学科重点项目(No.2010-2012);"十一五"国家科技支撑计划(No.2006BAI05A07)

摘　　要：目的:探索维甲酸(RA)体外诱导P19胚胎癌细胞向神经元样细胞分化过程中分化相关因子的表达。方法:小鼠P19胚胎癌细胞随机分为对照组和实验组,实验组予RA添加培养,对照组予同体积的无RA溶解液无水乙醇代替,诱导4 d后,分化7 d。通过免疫荧光法鉴定细胞分化,实时定量PCR和Western blot检测分化相关指标SOX2、NESTIN、TUJ1、Neurod2的表达。结果:分化7 d后,在实验组观察到丰富的TUJ1的表达;对照组极少观察到TUJ1表达。实验组的SOX2呈波动高表达,对照组持续递减表达;NESTIN呈现迅猛上升,在诱导第4天时明显高于对照组,分化期间两组无显著性差异;Neurod2在诱导第2天有小量表达,迅速升高至分化第1天出现峰值。对照组中Neurod2和TUJ1显著性低表达,在实验组中均高表达。结论:RA体外诱导P19胚胎癌细胞向神经元样细胞分化可作为研究人类神经系统发育的体外模型;Neurod2可能促进神经前体细胞向神经元的分化和维持神经功能活性。Objective： To explore the expression of differentiation related factors in the process of inducing P19 embryonic carcinoma cells differentiation to neural cells by retinoic acid （RA）. Methods： The P19 embryonic carcinoma cells were randomly divided into control group and experimental group. The experimental group was treated by RA while the control group was treated by equal volume anhydrous ethanol without RA. All the cells were induced for 4 days and differentiated for 7-days. The neural differentiation was determined using immuno-cytochemisty method. Real-time qPCR and Western blot were employed to analyse gene and protein changes ofSOX2, NESTIN, TUJ 1, Neurod2. Results： At day 7 after differentiation, abundant TUJ 1 expression was observed in the experimental group, compared with little expression in the control group. SOX2 was highly expressed in the experimental group while it progressively decreased in the control group. The expression of NESTIN increased rapidly which was significantly higher in the experimental group than that in the control group at day 4 after induction but was comparable without significant difference during differentiation. The expression of Neurod2 was observed at day 2 during induction which increased and peaked at d 1 during differentiation. Both Neurod2 and TUJ1 were highly expressed in the experimental group compared with the low expression in the control after RA induction. Conclusion： The RA-induced P19 embryonic carcinoma cell differentiation to neuron-like cells can be used as a vitro model for neuronal development and differentiation research.

关 键 词：Neurod2 维甲酸 P19胚胎癌细胞 神经元 诱导分化 

分 类 号：R741[医药卫生—神经病学与精神病学] R741.02[医药卫生—临床医学]